Kinetically controlled enzymatic synthesis of dipeptide precursor of <scp>l</scp>‐alanyl–<scp>l</scp>‐glutamine

Wang, Mengfan; Qi, Wei; Yu, Qingxin; Su, Rongxin; He, Zhimin

doi:10.1002/bab.55

Cited by 15 publications

(16 citation statements)

References 27 publications

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“…Reference source not found.C, the highest dipeptide yield was observed at 50 o C for both free PA and PA@MNCC, which was lower than the optimal temperature for PA activity in 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 21 BAEE hydrolysis in this study. Similar results were observed previously 13 . With an increase in temperature from 50 o C to 70 o C, the yield of Z-Ala-Gln catalyzed by free PA sharply decreased to about 15%, while the yield in the PA@MNCC-catalyzed reaction was more that 50%.…”

Section: Pa@mncc-catalyzed Biosynthesis Of Z-ala-gln In Dessupporting

confidence: 93%

“…Reference source not found.B, with increased KCl concentration the yield of Z-Ala-Gln catalyzed by free PA and/or PA@MNCC decreased by 18% and 8%, respectively. This decrease was mainly caused by high ionic strength which changed the affinity of PA to the Z-Ala-OMe substrate and led to a lower yield 13,59 . Moreover, it should be noted that the yield of PA@MNCC catalytic Z-Ala-Gln synthesis was approximately 58.6%, which was about 10% higher than the yield catalyzed by its free counterpart.…”

Section: Pa@mncc-catalyzed Biosynthesis Of Z-ala-gln In Desmentioning

confidence: 99%

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Papain@Magnetic Nanocrystalline Cellulose Nanobiocatalyst: A Highly Efficient Biocatalyst for Dipeptide Biosynthesis in Deep Eutectic Solvents

Cao

et al. 2015

ACS Sustainable Chem. Eng.

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Magnetic nanocrystalline cellulose (MNCC), a novel bio-based nanocomposite, was prepared via a simple co-precipitation-cross-linking technique and structurally characterized. Papain (PA) was successfully immobilized onto the MNCC. The resulting nano-biocatalyst PA@MNCC showed high PA loading (333 mg/g) and enzyme activity recovery (more than 80%). The stabilities of PA@MNCC was greatly superior to that of its free counterpart. Also, PA@MNCC manifested markedly enhanced solvent tolerance. The secondary structure study of the enzyme proved that these enhancements were attributed to the increase of structure rigidity of PA@MNCC. The observed optimum pH and temperature of PA@MNCC were significantly higher than the corresponding levels of free PA. A kinetic study demonstrated that PA@MNCC had a increase in enzyme-substrate affinity.Furthermore, the as-prepared PA@MNCC was successfully used as an efficient biocatalyst for the synthesis of N-(benzyloxycarbonyl)-alanyl-glutamine (Z-Ala-Gln) dipeptide in deep eutectic solvent (DES), choline chloride (ChCl):Urea(1:2), with a high yield (about 71.5%), which, to our knowledge, was greatly higher than that reported previously. Besides, the novel PA@MNCC was easily recycled from the reaction medium by magnetic forces. Obviously, MNCC is a promising and competitive enzyme carrier and the as-prepared nano-biocatalyst PA@MNCC exhibited great potential for efficient biosynthesis of dipeptide.

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Section: Pa@mncc-catalyzed Biosynthesis Of Z-ala-gln In Dessupporting

confidence: 93%

Section: Pa@mncc-catalyzed Biosynthesis Of Z-ala-gln In Desmentioning

confidence: 99%

Papain@Magnetic Nanocrystalline Cellulose Nanobiocatalyst: A Highly Efficient Biocatalyst for Dipeptide Biosynthesis in Deep Eutectic Solvents

Cao

et al. 2015

ACS Sustainable Chem. Eng.

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“…This steric problem will be coupled to the standard diffusional limitations, even if the active centre is fully intact [31] or a non-porous nanoparticle is utilized [32]. However, due to the multiple uses of proteases, this should not discard the biocatalysts for other uses, for example synthesis of peptide bonds [33][34][35]. Ficin has been immobilized in just some few examples [36][37][38][39][40].…”

Section: Introductionmentioning

confidence: 99%

Immobilization/Stabilization of Ficin Extract on Glutaraldehyde-Activated Agarose Beads. Variables That Control the Final Stability and Activity in Protein Hydrolyses

et al. 2018

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Ficin extract has been immobilized on different 4% aminated-agarose beads. Using just ion exchange, immobilization yield was poor and expressed activity did not surpass 10% of the offered enzyme, with no significant effects on enzyme stability. The treatment with glutaraldehyde of this ionically exchanged enzyme produced an almost full enzyme inactivation. Using aminated supports activated with glutaraldehyde, immobilization was optimal at pH 7 (at pH 5 immobilization yield was 80%, while at pH 9, the immobilized enzyme became inactivated). At pH 7, full immobilization was accomplished maintaining 40% activity versus a small synthetic substrate and 30% versus casein. Ficin stabilization upon immobilization could be observed but it depended on the inactivation pH and the substrate employed, suggesting the complexity of the mechanism of inactivation of the immobilized enzyme. The maximum enzyme loading on the support was determined to be around 70 mg/g. The loading has no significant effect on the enzyme stability or enzyme activity using the synthetic substrate but it had a significant effect on the activity using casein; the biocatalysts activity greatly decreased using more than 30 mg/g, suggesting that the near presence of other immobilized enzyme molecules may generate some steric hindrances for the casein hydrolysis.

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“…The commercial demand for AQ is increasing with the expansion of new applications and the development of new products based on AQ. Chemical and biotechnological methods for AQ production have been developed [12][13][14][15][16]. AQ can be synthesized by chemical condensation of benzyloxycarbonyl-l-alanine and methyl glutamate via activated ester [17] or triphenyl phosphine/ hexachloroethane condensation, which used to require complex steps such as amino acid activation, subunit protection, and removing the protective groups.…”

Section: Introductionmentioning

confidence: 99%

Metabolic engineering of Escherichia coli for efficient production of l-alanyl-l-glutamine

et al. 2020

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Background: l-Alanyl-l-glutamine (AQ) is a functional dipeptide with high water solubility, good thermal stability and high bioavailability. It is widely used in clinical treatment, post-operative rehabilitation, sports health care and other fields. AQ is mainly produced via chemical synthesis which is complicated, time-consuming, labor-intensive, and have a low yield accompanied with the generation of by-products. It is therefore highly desirable to develop an efficient biotechnological process for the industrial production of AQ. Results: A metabolically engineered E. coli strain for AQ production was developed by over-expressing l-amino acid α-ligase (BacD) from Bacillus subtilis, and inactivating the peptidases PepA, PepB, PepD, and PepN, as well as the dipeptide transport system Dpp. In order to use the more readily available substrate glutamic acid, a module for glutamine synthesis from glutamic acid was constructed by introducing glutamine synthetase (GlnA). Additionally, we knocked out glsA-glsB to block the first step in glutamine metabolism, and glnE-glnB involved in the ATP-dependent addition of AMP/UMP to a subunit of glutamine synthetase, which resulted in increased glutamine supply. Then the glutamine synthesis module was combined with the AQ synthesis module to develop the engineered strain that uses glutamic acid and alanine for AQ production. The expression of BacD and GlnA was further balanced to improve AQ production. Using the final engineered strain p15/AQ10 as a whole-cell biocatalyst, 71.7 mM AQ was produced with a productivity of 3.98 mM/h and conversion rate of 71.7%. Conclusion: A metabolically engineered strain for AQ production was successfully developed via inactivation of peptidases, screening of BacD, introduction of glutamine synthesis module, and balancing the glutamine and AQ synthesis modules to improve the yield of AQ. This work provides a microbial cell factory for efficient production of AQ with industrial potential.

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Kinetically controlled enzymatic synthesis of dipeptide precursor of l‐alanyl–l‐glutamine

Cited by 15 publications

References 27 publications

Papain@Magnetic Nanocrystalline Cellulose Nanobiocatalyst: A Highly Efficient Biocatalyst for Dipeptide Biosynthesis in Deep Eutectic Solvents

Papain@Magnetic Nanocrystalline Cellulose Nanobiocatalyst: A Highly Efficient Biocatalyst for Dipeptide Biosynthesis in Deep Eutectic Solvents

Immobilization/Stabilization of Ficin Extract on Glutaraldehyde-Activated Agarose Beads. Variables That Control the Final Stability and Activity in Protein Hydrolyses

Metabolic engineering of Escherichia coli for efficient production of l-alanyl-l-glutamine

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